Mechanisms of Ub-independent ODC recognition and translocation by the 26S proteasome

Mechanisms of Ub-independent ODC recognition and translocation by the 26S proteasome | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Biological Sciences - Article Mechanisms of Ub-independent ODC recognition and translocation by the 26S proteasome Yao Cong, kaijian chen, Yanxing Wang, Xingyan Ye, Zhanyu Ding, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8328438/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted You are reading this latest preprint version Abstract While most eukaryotic proteins require ubiquitination for proteasomal degradation, a considerable number of substrates undergo ubiquitin-independent degradation. Ornithine decarboxylase (ODC) is a well-known substrate degraded by the 26S proteasome in a ubiquitin- and cofactor-independent manner. However, the underling mechanism of how ODC is recognized and processed without the ubiquitin tag by the 26S proteasome remain unclear. Here, we present eleven cryo-EM structures of the human 26S proteasome in complex with C-terminal truncated or full-length ODC, capturing the entire degradation process from initial recognition to final unfolding. These structures reveal a dynamic, multivalent recognition process involving sequential engagement with vWA domain of Rpn10, Rpt4/5’s coiled-coil, and Rpn2’s PC domain, collectively bypassing the need for ubiquitin. Although the folded ODC core is sufficient for initial binding, its flexible C-terminal tail is crucial for allosterically activating the proteasome’s AAA+ ATPase motor. Furthermore, we also identify a unique translocation gateway where Rpn11’s JAMM motif guides the ODC C-tail into the AAA+ATPase ring, repurposing this deubiquitinase for a novel function. Once activated, the proteasome translocates ODC C-tail and unfolds ODC using its canonical ATP-driven machinery. The continuous structural series from initial substrate recognition through final unfolding elucidates the complete molecular mechanism of ubiquitin-independent ODC degradation. Biological sciences/Biochemistry/Proteolysis/Proteasome Biological sciences/Biochemistry/Structural biology/Electron microscopy/Cryoelectron microscopy Biological sciences/Biochemistry/Proteolysis/Protein quality control Cryo-EM Proteasome Ubiquitin-independent degradation ODC Rpn10 Rpn11 Full Text Additional Declarations There is NO Competing Interest. Supplementary Files validation.pdf cryo-EM structure model validation report 26SODC3DVA3.mp4 26S-ODCΔC-3DVA analysis Cite Share Download PDF Status: Under Review Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8328438","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Biological Sciences - Article","associatedPublications":[],"authors":[{"id":569244678,"identity":"3681d51f-3bf2-4fa5-b627-1cb2e980e3a7","order_by":0,"name":"Yao 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Ornithine decarboxylase (ODC) is a well-known substrate degraded by the 26S proteasome in a ubiquitin- and cofactor-independent manner. However, the underling mechanism of how ODC is recognized and processed without the ubiquitin tag by the 26S proteasome remain unclear. Here, we present eleven cryo-EM structures of the human 26S proteasome in complex with C-terminal truncated or full-length ODC, capturing the entire degradation process from initial recognition to final unfolding. These structures reveal a dynamic, multivalent recognition process involving sequential engagement with vWA domain of Rpn10, Rpt4/5’s coiled-coil, and Rpn2’s PC domain, collectively bypassing the need for ubiquitin. Although the folded ODC core is sufficient for initial binding, its flexible C-terminal tail is crucial for allosterically activating the proteasome’s AAA+ ATPase motor. 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